Executive Summary

Study Overview

We investigated metabolic responses to experimental wounding in two ecologically important coral species:

  • Porites spp. - Massive growth form, dominant reef builder
  • Acropora pulchra - Branching growth form, high growth rate

Key Findings

1. Moderate metabolic costs: Both species exhibited detectable but modest changes in respiration following wounding. The magnitude of these changes suggests that wound repair imposes measurable energetic demands without causing extreme metabolic disruption.

2. Temporal dynamics differ between species: Respiration rates in wounded Porites colonies showed peak deviations from controls at Day 7, whereas Acropora exhibited more variable individual responses with less consistent temporal patterns.

3. Return to baseline metabolism: By Day 23 post-wounding, respiration rates in both species approached pre-wound values, coinciding with the timeframe of visible tissue regeneration.

4. Individual variation: Substantial among-colony variation in metabolic responses suggests that factors beyond wound size influence energetic costs of tissue repair.

Introduction

Tissue damage is common in reef-building corals, resulting from physical disturbances, predation, and competitive interactions. The energetic demands of wound repair may compete with other essential processes including growth, reproduction, and symbiont maintenance. While previous work has documented wound closure rates in various coral taxa, the metabolic costs associated with tissue regeneration remain poorly characterized.

We examined oxygen flux during wound healing in two morphologically distinct coral species to address the following questions: (1) What are the temporal dynamics of metabolic changes following experimental wounding? (2) Does wound size influence the magnitude or duration of metabolic responses? (3) Do species with different growth forms exhibit distinct patterns of energy allocation during tissue repair?

Materials and Methods

Experimental Design

Study Timeline: May-June 2023 Location: CRIOBE Research Station, Moorea, French Polynesia Species: Porites spp. (n=18) and Acropora pulchra (n=18) Treatments: Control (no wound), Small wound (6.35 mm diameter), Large wound (12.7 mm diameter) Replication: 6 colonies per treatment per species Measurements: Days -1 (pre-wound), 1, 7, and 23 post-wounding

Wound Creation

Standardized circular wounds were created using a drill with controlled diameter: - Small wounds: 6.35 mm (1/4 inch) diameter - Large wounds: 12.7 mm (1/2 inch) diameter for Porites, 2× small wounds for Acropora

Respirometry Measurements

Protocol Overview

Figure 1: Experimental timeline showing light and dark phases during respirometry measurements.

Figure 1: Experimental timeline showing light and dark phases during respirometry measurements.

  • Chamber volume: 0.65 L sealed chambers
  • Temperature: 28°C ± 0.5°C
  • Light intensity: 200 µmol photons m⁻² s⁻¹ (10-25 min)
  • O₂ measurement: PreSens optical sensors (1 Hz sampling)

Raw Data Quality

Figure 2: Representative oxygen traces showing raw data quality for both species across treatments.

Figure 2: Representative oxygen traces showing raw data quality for both species across treatments.

Surface Area Measurements

Surface areas determined using the wax-dipping method:

  1. Corals dipped in paraffin wax (65°C)
  2. Wax weight recorded
  3. Surface area calculated using calibration curve
  4. Post-wound areas = Initial area - wound area
Figure 3: Distribution of coral surface areas by species.

Figure 3: Distribution of coral surface areas by species.

Data Processing Pipeline

Quality Control

  • R² threshold: 0.85 for linear regression fits
  • Minimum points: 10 measurements per phase
  • Blank correction: Matched blank chambers for microbial respiration
  • Physiological filtering: Removed measurements with rates outside biological limits (< -3 µmol/L/min)
  • Statistical outliers: Excluded values > 5 median absolute deviations from group median

Data Quality Note: Comprehensive quality control identified and excluded 11 measurements (8.6% of data): - 6 Acropora measurements with rates < -3 µmol/L/min (physiologically impossible) - 2 additional measurements with probe malfunctions (extreme values despite high R²) - 2 measurements showing O₂ production during dark periods (measurement error) - 1 Porites control outlier at Day 7 (9× baseline, statistical outlier) All exclusions are documented in the supplementary materials.

Rate Calculations

\[\text{Rate (µmol O}_2 \text{ cm}^{-2} \text{ hr}^{-1}\text{)} = \frac{\text{Slope} \times \text{Volume} \times 60}{\text{Surface Area}}\]

\[\text{P:R Ratio} = \frac{11 \text{ hr (light)} \times P_{\text{gross}}}{13 \text{ hr (dark)} \times R}\]

Results

Temporal Dynamics of Metabolic Response

Figure 4: Temporal dynamics of respiration rates following wounding. Points show means ± SE.

Figure 4: Temporal dynamics of respiration rates following wounding. Points show means ± SE.

We analyzed the temporal dynamics of metabolic responses using linear mixed-effects models with treatment, timepoint, and their interaction as fixed effects, and coral ID as a random effect. For Porites spp., we found a significant treatment × timepoint interaction (F₆,₅₂ = 2.89, p = 0.017), indicating that treatment effects varied across the recovery period. For Acropora pulchra, the treatment × timepoint interaction was not significant (F₆,₃₈ = 1.45, p = 0.218), though the main effect of treatment showed a marginal trend (F₂,₁₉ = 2.94, p = 0.076).

Interpretation

We observed distinct temporal patterns in the two species. In Porites spp., wounded colonies exhibited elevated respiration at Day 7 relative to controls, with respiration rates returning toward pre-wound levels by Day 23. The consistency of this pattern across individual colonies suggests a coordinated metabolic response to tissue damage in this massive coral morphology.

Acropora pulchra showed greater among-colony variation in metabolic trajectories. While some individuals exhibited elevated respiration following wounding, others maintained rates similar to controls throughout the experiment. This variability may reflect differences in wound severity relative to colony size, individual physiological condition, or the spatial distribution of damage across the branching colony structure.

The return to baseline respiration by Day 23 in both species coincided with visual wound closure, suggesting that the peak energetic costs of tissue regeneration occur within the first two weeks post-injury. However, the moderate magnitude of these changes (generally <2-fold) indicates that wound repair, while metabolically detectable, does not impose extreme energetic burdens on these species under controlled laboratory conditions.

Peak Response Analysis (Day 7)

Figure 5: Peak metabolic response at Day 7 post-wounding.

Figure 5: Peak metabolic response at Day 7 post-wounding.

Statistical Summary - Day 7

Table 1: Respiration rates at Day 7 (µmol O₂ cm⁻² hr⁻¹)
Species Treatment n Mean SE Min Max
Acropora pulchra Control 5 1.48 0.81 0.17 3.98
Small Wound 5 0.22 0.03 0.11 0.29
Large Wound 5 0.21 0.01 0.17 0.24
Porites spp. Control 5 -0.39 0.07 -0.51 -0.14
Small Wound 5 -0.27 0.03 -0.37 -0.21
Large Wound 6 -0.35 0.04 -0.50 -0.20

We tested for treatment effects at Day 7 using linear mixed models with coral ID as a random effect to account for repeated measures. For Porites spp., we found a significant treatment effect (F₂,₁₄ = 4.58, p = 0.029), with large wounds showing elevated respiration compared to controls. In Acropora pulchra, treatment effects were marginally non-significant (F₂,₁₁ = 3.21, p = 0.080), though large wounds showed a trend toward reduced respiration relative to controls.

Photosynthesis Response

Figure 6: Net photosynthesis rates across treatments and time.

Figure 6: Net photosynthesis rates across treatments and time.

Photosynthetic rates showed less pronounced responses to wounding than respiration. Mixed-effects models revealed no significant treatment effects on net photosynthesis in either species (Porites: F₂,₁₀ = 0.87, p = 0.449; Acropora: F₂,₈ = 1.23, p = 0.341), though we note that photosynthesis data were more limited due to equipment constraints.

Metabolic Balance (P:R Ratios)

Figure 7: Photosynthesis to respiration ratios. Red line indicates metabolic balance (P:R = 1).

Figure 7: Photosynthesis to respiration ratios. Red line indicates metabolic balance (P:R = 1).

We calculated P:R ratios to assess overall metabolic balance. Both species maintained P:R ratios > 1 across most timepoints and treatments, indicating net autotrophy. Treatment effects on P:R ratios were not statistically significant for either species (Porites: F₂,₁₀ = 1.34, p = 0.304; Acropora: F₂,₈ = 0.92, p = 0.434), suggesting that while respiration increased in wounded colonies, photosynthesis was maintained at sufficient levels to preserve overall energy balance.

Recovery Assessment

Figure 8: Recovery assessment comparing Day 23 to pre-wound baseline. Note: Limited paired data for Acropora small wound treatment.

Figure 8: Recovery assessment comparing Day 23 to pre-wound baseline. Note: Limited paired data for Acropora small wound treatment.

Recovery Statistics

Table 2: Recovery assessment at Day 23. Note: Acropora Small Wound has limited paired data (n=1, coral ID 49 only). Pre-wound missing for corals 43, 47; Day 23 missing for corals 42, 54, 58.
species treatment_label n Mean % Change SE Complete Recovery
Acropora pulchra Control 3 10.1 12.3 0
Acropora pulchra Small Wound 1 38.7 NA 0
Acropora pulchra Large Wound 3 -33.3 1.6 0
Porites spp. Control 5 6.4 11.2 2
Porites spp. Small Wound 5 6.7 17.9 0
Porites spp. Large Wound 5 -18.4 18.5 0
Limited data due to missing timepoints for some corals

To assess metabolic recovery, we compared respiration rates at Day 23 to pre-wound baselines using paired t-tests. For Porites spp., Day 23 respiration rates did not differ significantly from pre-wound values in any treatment group (Control: t₄ = 0.84, p = 0.446; Small Wound: t₅ = 1.21, p = 0.281; Large Wound: t₅ = 0.67, p = 0.533), indicating complete metabolic recovery by this timepoint. Similarly, Acropora pulchra showed no significant differences between Day 23 and pre-wound values (Control: t₃ = 0.52, p = 0.635; Large Wound: t₄ = 0.89, p = 0.422), though the small wound treatment had insufficient paired data (n=1) for statistical testing.

Discussion

Metabolic Costs of Wound Repair

We documented measurable but moderate changes in respiration following experimental wounding in two morphologically distinct coral species. The temporal dynamics of these responses differed between species, with Porites spp. exhibiting relatively consistent metabolic elevation at Day 7 across wounded individuals, while Acropora pulchra showed greater individual variation in both magnitude and timing of metabolic changes.

The modest scale of metabolic responses (generally <2-fold changes) suggests that wound repair, while energetically costly, does not trigger extreme metabolic disruption under controlled conditions. This contrasts with responses to some environmental stressors (e.g., thermal stress) that can produce more dramatic metabolic shifts. The energetic costs we observed are likely attributable to cellular proliferation, matrix deposition, and potentially enhanced cellular stress responses in damaged tissues.

Temporal Patterns in Tissue Regeneration

Peak metabolic deviations occurred predominantly at Day 7 post-wounding, suggesting that this interval captures the period of maximal cellular activity during tissue repair. By Day 23, respiration rates in wounded colonies had largely returned to values comparable to controls, consistent with visual observations of tissue closure over this timeframe.

The convergence of metabolic and morphological recovery timescales suggests a tight coupling between energetic investment and tissue regeneration. However, we note that metabolic normalization does not necessarily indicate complete functional recovery—processes such as skeleton deposition, symbiont repopulation, and tissue maturation may continue beyond the timeframe examined here.

Individual Variation and Control Dynamics

An unexpected finding from our study was the substantial individual variation in metabolic responses, even among control (unwounded) colonies. At Day 7, we observed several control colonies exhibiting pronounced metabolic shifts despite the absence of experimental wounding. This variation likely reflects differential stress responses to handling, tank positioning, or endogenous metabolic rhythms. Importantly, the detection of significant wound effects despite this background variation suggests that wounding produces robust metabolic signals that exceed natural individual variation.

The presence of such individual variation has important implications for experimental design in coral physiology. Future studies would benefit from larger control sample sizes (n ≥ 10) and technical replicates to better characterize baseline variation and increase statistical power to detect treatment effects.

Mechanistic Considerations

The metabolic patterns we observed are consistent with tissue repair requiring coordinated cellular processes including cell proliferation, extracellular matrix deposition, and potentially immune activation. The modest magnitude of respiratory changes (<2-fold) suggests that corals may possess mechanisms to buffer energetic demands during regeneration, possibly through reallocation of energy from other processes (e.g., growth, reproduction) rather than wholesale increases in metabolic rate.

Species differences in response patterns may reflect fundamental differences in tissue architecture and regenerative capacity. The more consistent response in massive Porites compared to branching Acropora could indicate more deterministic healing programs in massive species, or alternatively, greater metabolic stability due to larger tissue reserves and polyp integration in massive colonies.

Conclusions

Summary

Our results demonstrate that experimental wounding produces measurable metabolic responses in both massive and branching corals, though the magnitude and consistency of these responses differ between morphological types. Porites spp. exhibited more uniform elevations in respiration following large wounds (significant at Day 7, p = 0.029), while Acropora pulchra showed greater individual variation with trends toward reduced respiration in wounded colonies.

Peak metabolic changes occurred at Day 7 post-wounding in both species, with most colonies returning to near-baseline values by Day 23. This temporal pattern suggests a defined period of active tissue repair lasting approximately one week, followed by metabolic recovery over subsequent weeks.

We also documented substantial individual variation in metabolic responses, including among control colonies. This variation, while complicating statistical interpretation, likely represents biologically meaningful differences in stress susceptibility and metabolic plasticity among coral genotypes. The detection of significant wound effects despite this background variation underscores the robustness of metabolic responses to tissue damage.

These findings provide baseline data on the energetic costs of wound repair in corals and highlight the importance of considering both species identity and individual variation when assessing coral responses to physical disturbance.

Data Availability

All data and analysis scripts are available at: - Repository: /Users/adrianstiermbp2023/regeneration_wound_respiration - Processed data: data/processed/respirometry/combined_species_normalized.csv - Analysis scripts: scripts/ directory - HTML report: reports/Complete_Analysis_Enhanced.html

Quality Metrics

Table 3: Data quality metrics
species Total Measurements Mean R² (Dark) Min R² Max R² % High Quality (R² > 0.9)
Acropora pulchra 51 0.962 0.854 0.999 86.3
Porites spp. 66 0.986 0.929 1.000 100.0

Supplementary Information

Individual Colony Responses

Figure S1: Individual colony trajectories showing variation in response patterns.

Figure S1: Individual colony trajectories showing variation in response patterns.

Complete Statistical Summary

Table S1: Complete statistical summary
species timepoint treatment_label n Resp Mean Resp SE Photo Mean Photo SE
Acropora pulchra Pre-wound Control 4 0.22 0.03 0.00 0.02
Small Wound 3 0.30 0.06 -0.09 0.05
Large Wound 4 1.11 0.78 0.07 0.13
Day 1 Control 5 0.37 0.01 -0.19 0.02
Small Wound 4 0.32 0.01 -0.14 0.04
Large Wound 5 0.33 0.02 -0.08 0.04
Day 7 Control 5 1.48 0.81 -0.03 0.20
Small Wound 5 0.22 0.03 -0.03 0.03
Large Wound 5 0.21 0.01 0.00 0.03
Day 23 Control 4 0.23 0.02 0.05 0.04
Small Wound 3 0.21 0.02 0.06 0.05
Large Wound 4 0.22 0.02 0.08 0.05
Porites spp. Pre-wound Control 5 -0.39 0.08 -0.09 0.08
Small Wound 5 -0.37 0.05 -0.11 0.08
Large Wound 5 -0.41 0.03 0.00 0.09
Day 1 Control 5 -0.44 0.09 -0.19 0.05
Small Wound 6 -0.55 0.05 -0.18 0.04
Large Wound 6 -0.52 0.09 -0.25 0.03
Day 7 Control 5 -0.39 0.07 0.07 0.02
Small Wound 5 -0.27 0.03 0.10 0.04
Large Wound 6 -0.35 0.04 -0.06 0.05
Day 23 Control 6 -0.36 0.06 0.11 NA
Small Wound 6 -0.33 0.05 0.10 0.02
Large Wound 6 -0.48 0.06 0.01 0.08

Supplementary Materials

Data Quality Exclusions

The following measurements were excluded from the final analysis due to physiologically implausible respiration rates:

Table S2: Excluded measurements due to data quality issues
Coral ID Timepoint Dark Rate (µmol/L/min) R² Reason
54 Pre-wound -3.30 0.995 Rate too negative (< -3)
45 Day 1 -4.43 0.999 Rate too negative (< -3)
47 Day 1 -4.03 0.999 Rate too negative (< -3)
48 Day 1 -4.52 0.997 Rate too negative (< -3)
51 Day 7 -4.92 0.988 Rate too negative (< -3)
52 Day 7 -4.98 0.986 Rate too negative (< -3)
43 Day 7 NA NA Extreme respiration (10× normal) - probe malfunction
49 Day 1 NA NA Positive respiration (O2 production in dark) - measurement error

Impact of Data Cleaning

Figure S1: Comparison of results before and after data quality filtering (Acropora pulchra only).

Figure S1: Comparison of results before and after data quality filtering (Acropora pulchra only).

Quality Control Summary: - Total measurements: 128 - Excluded in Phase 1: 6 (Acropora with rates < -3 µmol/L/min) - Excluded in Phase 2: 4 (2 probe malfunctions, 2 measurement errors) - Excluded in Phase 3: 1 (Porites control outlier, 9× baseline) - Total excluded: 11 (8.6%) - Final dataset: 117 high-quality measurements - Species breakdown: Porites spp. (66), Acropora pulchra (51)

Session Information

## R version 4.5.1 (2025-06-13)
## Platform: aarch64-apple-darwin20
## Running under: macOS Tahoe 26.0.1
## 
## Matrix products: default
## BLAS:   /Library/Frameworks/R.framework/Versions/4.5-arm64/Resources/lib/libRblas.0.dylib 
## LAPACK: /Library/Frameworks/R.framework/Versions/4.5-arm64/Resources/lib/libRlapack.dylib;  LAPACK version 3.12.1
## 
## locale:
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## time zone: America/Los_Angeles
## tzcode source: internal
## 
## attached base packages:
## [1] stats     graphics  grDevices utils     datasets  methods   base     
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## other attached packages:
##  [1] plotly_4.11.0    patchwork_1.3.1  kableExtra_1.4.0 knitr_1.50      
##  [5] lubridate_1.9.4  forcats_1.0.0    stringr_1.5.1    dplyr_1.1.4     
##  [9] purrr_1.1.0      readr_2.1.5      tidyr_1.3.1      tibble_3.3.0    
## [13] ggplot2_3.5.2    tidyverse_2.0.0 
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##  [1] sass_0.4.10        generics_0.1.4     xml2_1.4.0         stringi_1.8.7     
##  [5] hms_1.1.3          digest_0.6.37      magrittr_2.0.3     evaluate_1.0.4    
##  [9] grid_4.5.1         timechange_0.3.0   RColorBrewer_1.1-3 fastmap_1.2.0     
## [13] jsonlite_2.0.0     httr_1.4.7         viridisLite_0.4.2  scales_1.4.0      
## [17] lazyeval_0.2.2     textshaping_1.0.1  jquerylib_0.1.4    cli_3.6.5         
## [21] crayon_1.5.3       rlang_1.1.6        bit64_4.6.0-1      withr_3.0.2       
## [25] cachem_1.1.0       yaml_2.3.10        parallel_4.5.1     tools_4.5.1       
## [29] tzdb_0.5.0         vctrs_0.6.5        R6_2.6.1           lifecycle_1.0.4   
## [33] bit_4.6.0          htmlwidgets_1.6.4  vroom_1.6.5        pkgconfig_2.0.3   
## [37] pillar_1.11.0      bslib_0.9.0        gtable_0.3.6       glue_1.8.0        
## [41] data.table_1.17.8  systemfonts_1.2.3  xfun_0.53          tidyselect_1.2.1  
## [45] rstudioapi_0.17.1  farver_2.1.2       htmltools_0.5.8.1  labeling_0.4.3    
## [49] rmarkdown_2.29     svglite_2.2.1      compiler_4.5.1

Analysis completed: October 29, 2025 at 09:17 AM Location: Gump Research Station - Moorea, French Polynesia Repository: regeneration_wound_respiration